The gastrointestinal mucosa maintains its integrity in the presence of powerful digestive enzyme systems capable of breaking down proteins, fats and carbohydrates. The intestines themselves are composed of these very molecules, yet they resist digestion. Occasionally, molecular defense mechanisms fail and mucosal breakdown occurs. Examples of such failure are duodenal ulcers, gastric ulcers, and peptic esophagitis. Hundreds of thousands of Americans suffer morbidity and also significant mortality from these conditions every year. The purpose of this project will be to study pathophysiologic mechanisms of gastric mucosal injury with the ultimate goal of developing new therapeutic strategies. The specific aims of the work to be accomplished in this project are to explore the role of calcium in gastric mucosal injury. Cells normally maintain a low intracellular free calcium concentration in the range of 100 nM. They maintain this low intracellular calcium concentration of 1-2mM calcium. When cells are injured, they lose the ability to maintain this calcium concentration gradient. We hypothesize that disruption of the homeostatic mechanisms for controlling intracellular calcium concentration is an early and critical step in cell injury. We propose to utilize FURA-2 to study intracellular calcium homeostasis in cells of the gastric mucosa of rabbits. We will study the effect of pH, temperature, bile acids, hypoxia, and free radicals on calcium homeostasis. We will explore the possibility that calcium entry into the cell is part of the pathophysiology of injury rather than just a result of injury. To investigate this possibility, we will compare the time of onset as well as the threshold of injury that raises intracellular calcium as opposed to those damaging other aspects of cellular function such as oxygen consumption, Trypan blue exclusion, or morphology. We will also determine whether injury-induced intracellular hypercalcemia is reversible. If it is reversible, this would suggest that it is certainly not a terminal event for the cell. We will use two basic approaches in our studies. One will be a mixed-cell approach in which cells are studied in suspension using a spectrofluorometer. The second approach will be to study gastric mucosal cells individually utilizing spectrofluorescent video photomicroscopy.